The goal of our first cycle of funding was to examine the molecular mediators of tissue interactions that regulate craniofacial development. We focused on the role that the brain plays in establishing a signaling center, the Frontonasal Ectodermal Zone (FEZ), because the FEZ regulates patterned growth of the upper jaw. Our research revealed that the brain controls the spatial organization of the FEZ, which then produces unique facial morphologies including disease phenotypes that fall along a continuous spectrum of morphologies. Our preliminary data suggest a novel mechanism by which the brain controls the spatial organization of the FEZ. These data also indicate that the FEZ is dynamic and shifts expression boundaries to regulate multiple morphogenetic processes during facial development. Finally, cellular responses to Fgf signaling during facial development have been largely unknown, and our preliminary studies suggest that basic cellular processes are controlled by FGF signaling. In response to FGF signaling these cellular behaviors participate in regulating facial morphogenesis. Collectively, these data suggest mechanisms that are used for development, integration, and fusion of facial primordia. Based on these data, we hypothesize that SHH from the brain controls facial morphogenesis by inducing a negative feedback loop in the FEZ via microRNAs and this helps to define the domain of Shh in the FEZ. The FEZ then orchestrates its own changing expression domain to control morphogenetic processes such as proliferation, migration, and polarization of mesenchymal cells that drive growth and fusion of the facial primordia that form the upper jaw. We will test this hypothesis in three Specific Aims. First, we will examine the extent to which these micro RNAs regulate the spatial organization of Shh expression in the FEZ and validate putative target genes. Second, the FEZ changes shape over time, and we will determine the role that novel expression domains of Shh play during growth and fusion of the median nasal, lateral nasal, and maxillary processes. Third, we will examine the role that downstream targets of FGF signaling have in mediating the cellular response to signaling by this pathway, and we will examine a mouse model of Crouzon Syndrome. The results from this research will lead to a more comprehensive understanding of mechanisms that control facial development and will identify putative disease causing mechanisms related to cleft lip and palate among other structural disease.